Development and application of technology platform for identification of the hist

描述（由申请人提供）：组蛋白n端尾部的翻译后修饰在表观遗传调控中起重要作用。组蛋白编码假说提出，组蛋白尾部的修饰产生组合结合位点，以募集酶进行转录调节。组蛋白修饰的失调与包括癌症在内的多种疾病有关。虽然对组蛋白修饰和修饰酶有很多了解，但修饰残基如何传递信息以调节基因表达仍不清楚。修饰特异性结合蛋白的鉴定将是填补这一知识空白的第一步。Lys16 （H4K16ac）乙酰化缺失和H4 (H4K20me3) Lys20三甲基化缺失与癌症相关，但分子机制尚不清楚。H4K16ac和H4K20me3的“结合物”尚不清楚，这阻碍了我们对这些修饰的缺失如何导致癌症的理解。我们建议首先开发一个蛋白质组学平台来识别特定的组蛋白修饰结合物，然后使用该平台来识别结合H4K16ac和H4K20me3的蛋白质。我们的蛋白质组学平台包括高效的蛋白质亲和纯化，修饰组蛋白肽与SILAC标记和纳米高效液相色谱-质谱/质谱用于蛋白质鉴定和定量。我们有初步的数据证明了使用在Lys4和/或Lys9位点进行修饰的组蛋白H3肽建立这样一个平台的可行性。该I期提案将允许我们：(1)开发一个技术平台，使我们能够识别和量化与组蛋白上任何修饰结合的蛋白质；(2)鉴定与H4K16ac和H3K20me3结合的可能具有肿瘤抑制功能的蛋白。我们的商业活动将包括：(1)为学术和工业实验室提供鉴定组蛋白编码效应器的合同服务；(2)开发组蛋白代码读取工具包(r)，该工具包由我们发现的与修饰组蛋白结合的重组蛋白和抗体组成，以促进表观遗传学领域组蛋白代码的功能分析。

DESCRIPTION (provided by applicant): Post-translational modifications of the N-terminal tail of histones play important roles in epigenetic regulation. The histone code hypothesis proposes that modifications of histone tails create combinatorial binding sites to recruit enzymes for transcriptional regulation. Dysregulation of histone modifications has been linked with diverse diseases including cancer. While much is known about histone modifications and the modifying enzymes, it remain largely unclear how the modified residues transmit the information to regulate gene expression. Identifications of modification-specific binding proteins will be the first step to fill this knowledge gap. Loss of acetylation of Lys16 (H4K16ac) and trimethylation of Lys20 of H4 (H4K20me3) has been correlated with cancer, but the molecular mechanism is not clear. The "binders" to H4K16ac and H4K20me3 are not known, hindering our understanding of how loss of these modifications may contribute to cancer. We propose to first develop a proteomic platform to identify specific histone modification binders and then use the platform to identify proteins that bind H4K16ac and H4K20me3. Our proteomics platform consists of efficient protein affinity purification with modified histone peptides with SILAC labeling and nanoHPLC-MS/MS for protein identification and quantification. We have preliminary data demonstrating the feasibility of such a platform using histone H3 peptides with modifications at Lys4 and/or Lys9. This Phase I proposal will allow us to: (1) develop a technology platform that will enable us to identify and quantify proteins that bind to any modifications on histone proteins; and (2) identify proteins that bind to H4K16ac and H3K20me3, that may have tumor suppressor functions. Our commercial activities will include: (1) contract services to academic and industry labs for the identification of histone code effectors; and (2) development of a Histone Code Reader Toolkit(r), which consists of recombinant proteins and antibodies that we find to bind modified histones to facilitate functional analysis of the histone code in epigenetics field.